RNA干扰p21联合顺铂对U2OS细胞增殖及凋亡的影响

Effect of RNA interference targeting p21 combined with cisplatin treatment on proliferation and apoptosis of U2OS cells

目的探讨RNA干扰siRNA联合顺铂对骨髓瘤细胞U2OS增殖和凋亡的影响及相关作用机制。方法将化学合成的针对p21的siRNA序列转染至U2OS细胞,分为5组:空白对照组、阴性对照组、干扰组、顺铂组、联合组(干扰+顺铂)。采用MTT比色法、流式细胞术和分光光度法分别检测U2OS细胞增殖、细胞凋亡、细胞周期及半胱氨酸天冬氨酸蛋白酶家族成员Caspase-9、Caspase-3和Caspase-6的活化程度。结果 p21siRNA转染后,相对于顺铂处理组,联合组细胞死亡率明显升高(P<0.01);细胞周期各时相重新分布,G0/G1期细胞显著减少(P<0.01),S期和G2/M期细胞则明显增多(P<0.01);Caspase-9、Caspase-3和Caspase-6的活化程度显著升高(P<0.01)。结论 p21靶向RNA干扰联合顺铂处理通过调节细胞周期进程而促进了细胞凋亡,p21可作为骨髓瘤基因治疗的后选新靶点。

Objective To explore the effect of RNA interference （RNAi） targeting p21 combined with cisplatin treatment on proliferation and apoptosis of U2OS cells and its related mechanism. Methods The siRNA sequence targeting p21 synthesized by chemical meth- od was transfected into U2OS cells. The U2OS cells were divided into five groups： blank control, negative control, siRNA, eisplatin treated and combination groups （siRNA combined with cisplatin treatment）. Then MTT assay, flow cytomctry and spectrophotometry were employed to detect variations of cell viability, apoptosis, cell cycle and Caspase-9, Caspase-3 and Caspase-6 activity. Results After transfection, compared to the cisplatin treated group, the cell death rate of the combination group was markedly increased （P 〈0.01 ）. Cell cycle phases were redistributed, cell number of G0/G, were significantly decreased （P 〈0. 01 ） , but cell number of S and G2/M cells were significantly increased （P 〈 0. 01 ） ; and activities of Caspase-9, Caspase-3 and Caspase-6 were obviously induced （P 〈 0. 01 ）. Conclusions siRNA combined with cisplatin treatment can induce cell apoptosis via regulating cell cycle process, p21 gene may be a candidate target in the gene therapy of osteocarcinoma.